The goal of the current application is to continue to characterize mu and kappa opioid receptor subtypes and to integrate our understanding of the molecular biology of the cloned opioid receptor families with their pharmacology. The concept of multiple classes of opioid receptors, proposed from clinical studies done 30 years ago, has become increasingly important in understanding opioid actions and in the development of nonabuseable analgesics. For years, investigations have focused upon selective drugs in the elucidation of opioid receptor subtypes. Using these approaches, several subtypes of mu and kappa receptors have been proposed, each with its own binding profile and pharmacological actions. Cell lines have been identified which contain many of these subtypes, including two human neuroblastoma cell lines which contain a kappa receptor subtype we recently proposed, kappa3. Studies using these cell lines have established different mechanisms through which all these subtypes inhibit cAMP accumulation, consistent with their distinct pharmacological profiles in vivo. The cloning of four opioid receptor gene families has now opened new opportunities in correlating the pharmacology with he molecular biology of these systems. Antisense studies in which oligodeoxynucleotides complementary to the mRNA encoding a specific receptor are administered to cells or in vivo have established the importance of the MOR-1 clone in morphine analgesia, the DOR-1 clone in delta analgesia and the KOR-1 clone in kappa1 analgesia. The recently cloned K)R-3 clone appears to be associated with kappa3 analgesia. We will continue our study of the receptors in neuroblastoma cell lines, but we also plan to explore the expression and function of KOR-3 in non-neuronal cells, including immune cells and an osteoblast-like cell (MC3T3-E1). Recent work from our laboratory has established the presence of KOR-3 in several B-cell lymphoma cell lines, including Raji and NBV. We will compare the receptor and its functions in these different classes of cells with results from the brain. Studies correlating the cloned receptors with opioid pharmacology have raised the possibility of splice variants and a novel receptor for morphine-6beta-glucuronide. We also propose to explore the possibility of these splice variants and their potential role in opioid pharmacology.